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BCRC Number: 80392
Organism: Chlorobaculum tepidum
Author: (Wahlund et al.) Imhoff
History: << NBRC << DSM << M. T. Madigan, TLS
Other Collection No.: NBRC 103806; DSM 12025; ATCC 49652; CIP 105973
Source: Acidic high sulfide hot springs
Isolated Info:
    Isolated location: Travelodge Stream, Sulphur Bay, North Island, New Zealand
Growth Conditions: 47°C 30 day(s)
Biosafety Level: 1
Oxygen Requirement: Anaerobic
Type Strain: YES
    Reference: B4502
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Imhoff, J. F. 2003. Phylogenetic taxonomy of the family Chlorobiaceae on the basis of 16S rRNA and fmo (Fenna-Matthews-Olson protein) gene sequences. Int. J. Syst. Evol. Microbiol. 53: 941-951 .
Wahlund, T.M., C. R. Woese, R. W. Castenholz ,and M. T. Madigan 1991. A thermophilic green sulfur bacterium from New Zealand hot springs, Chlorobium tepidum sp. nov. Arch. Microbiol. 156: 81-90.


Green Sulfur Bacterium Medium
Solution I*45 ml
Solution II**2.5 ml
Neutralized sulfide solution***2 ml
Acetate solution****0.5 ml
After aseptically inject solution II, sulfide solution and acetate solution into vessel of solution I, the pH is adjusted to 6.8 by adding of sterile 0.6 N H2SO4 with a syringe without opening the vessel. If it is necessary, 0.5 ml thiosulfate solution***** is injected.
*Solution I
KH2PO40.5 g
MgSO4·7H2O0.5 g
CaCl2·2H2O0.05 g
NH4Cl0.5 g
Ammonium acetate0.5 g
Ferric citrate solution (0.1%)5 ml
Vitamin B12 solution (0.001%)2 ml
Trace element solution SL-62 ml
Resazurin1 mg
Distilled water950 ml
Mix ingredients except vitamin solution, and distribute the medium into culture vessels under a stream of nitrogen gas and seal with butyl rubber stoppers. After autoclaving (121°C, 15 min), aseptically inject vitamin solution.

**Solution II
NaHCO31.5 g
Distilled water50 ml
pH8.2, Filter sterile

*** Neutralized sulfide solution
Na2S·9H2O1.5 g
Distilled water100 ml
The sulfide solution is prepared in a vessel sealed with a butyl rubber stopper. Autoclave at 121℃ for 15 min under a N2 atmosphere. After cooling to room temperature, the pH is adjusted to about 7.3 by adding of sterile 2 N H2SO4 drop-wise with a syringe without opening the bottle. Appearance of yellow colour indicates the pH is about 8. The vessel should be shaken continuously to avoid precipitation of elemental sulfur. The final solution should be transparent yellow.

****Acetate solution
Ammonium acetate2.5 g
Magnesium acetate2.5 g
Fructose2.5 g
Distilled water100 ml

*****Thiosulfate solution
Na2S2O3·5H2O5 g
Distilled water100 ml
Solution is prepared in a vessel sealed with a butyl rubber stopper. Autoclave at 121℃ for 15 min under a N2 atmosphere.

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